PO.PS01.01 · 人群科学
使用ONT/ELIGOS测序区分KRAS密码子12突变热点处的表观遗传标记和DNA加合物
Differentiating epigenetic marks and DNA adducts at the KRAS codon 12 mutation hotspot using ONT/ELIGOS sequencing
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作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
突变——无论是遗传获得还是后天获得——都是癌症发展的核心，减少突变发生是降低癌症发病率的有效策略。最简单地说，DNA突变源于易出错的DNA复制，或源于DNA损伤被错误修复后的DNA复制。从机制上讲，当聚合酶错误地复制DNA时会发生突变，而在存在DNA修饰或DNA加合物的情况下，错误率会显著增加。在DNA修饰中，诸如5-甲基-脱氧胞苷（5mdC）等表观遗传标记最为丰富，其频率约为每100个胞嘧啶中有一个甲基化胞嘧啶。这大大高于内源性DNA加合物（约每10,000个核苷酸中有一个）和外源性DNA加合物（通常低于每1亿个正常核苷酸中有一个）的频率。因此，仍不清楚某个特定突变是源于表观遗传过程中的错误，还是源于内源性和暴露诱导的DNA加合物。遗憾的是，用于DNA加合物检测的化学特异性方法往往无法提供关于其在基因组内位置的信息，从而无法识别导致特定突变的DNA加合物。为弥补这一知识空白，我们团队开发了一种基于纳米孔的DNA测序方法，称为“ELIGOS”（Oxford Nanopore Technology-Epitranscriptional/Epigenomical Landscape Inferring from Glitches of ONT Signals，牛津纳米孔技术——从ONT信号异常推断表观转录组/表观基因组图谱），该方法能够在对DNA进行测序的同时，以碱基对分辨率（i）检测并（ii）区分各种类型的表观遗传标记和暴露诱导的DNA加合物。ONT/ELIGOS提供了一种创新工具，用于研究预先选定的DNA区域中的DNA损伤和突变。在本工作中，我们展示了最新的ONT/ELIGOS研究结果，证明能够检测和识别KRAS癌症驱动基因内密码子12处的DNA加合物。除常见的表观遗传标记和暴露来源的DNA加合物外，我们目前的DNA加合物panel还包括常见的内源性损伤。这些新方法共同能够揭示单个DNA分子中的动态变化。
查看英文原文 English abstract
Mutations-whether inherited or acquired-are central to cancer development, and reducing mutagenesis is an effective strategy for lowering cancer rates. At its simplest, DNA mutations arise from error-prone DNA replication or from the replication of DNA following incorrect repair of DNA lesions. Mechanistically, mutations occur when polymerases erroneously replicate DNA, with error rates increasing significantly in the presence of DNA modifications or DNA adducts. Among DNA modifications, epigenetic marks such as 5-methyl-deoxycytidine (5mdC) are the most abundant, occurring at a frequency of approximately one methylated cytosine per 100 cytosines. This is substantially higher than the frequency of endogenous DNA adducts (about one in 10,000 nucleotides) and exogenous DNA adducts (often less than one in 100 million normal nucleotides). Therefore, it remains unclear whether a specific mutation arises from errors in epigenetic processes, or from endogenous- and exposure-induced DNA adducts. Chemically specific methods for DNA adduct detection, unfortunately, often do not provide information about their position within the genome, prohibiting identification of the DNA adduct causing specific mutations. To address these knowledge gap, our group developed a nanopore-based DNA sequencing approach, called “ELIGOS” (Oxford Nanopore Technology-Epitranscriptional/ Epigenomical Landscape Inferring from Glitches of ONT Signals) that can (i) detect and (ii) differentiate between various types of epigenic marks and exposure-induced DNA adducts at base pair resolution while sequencing the DNA. ONT/ELIGOS provides an innovative tool to study the DNA lesions and mutations in pre-selected DNA regions. In this work, we present our latest ONT/ELIGOS findings demonstrating the detection and identification of DNA adducts at codon 12 within the KRAS cancer driver gene. In addition to common epigenetic marks and exposure-derived DNA adducts, our current panel of DNA adducts includes common endogenous lesions. Together these novel approaches can reveal the dynamics changes in single DNA molecules.
利益披露 Disclosure
G. Boysen, None..
A. Kurilung, None..
V. Wanchai, None..
I. Nookaew, None..
S. Shuck, None.